Nociception is the process whereby primary afferent somatosensory neurons recognize and respond to noxious stimuli, resulting in pain and neurogenic inflammation. Members of the TRP ion channel family play important roles in nociception and pain by functioning as sensors for a variety of noxious stimuli, including heat, cold, and inflammatory agents. More broadly, genetic studies have highlighted the importance of these and other TRP channel subtypes in processes ranging from calcium adsorption to neuronal growth cone guidance, keratinocyte development, and numerous aspects of sensory transduction. Thus, understanding how these channels respond to physiological stimuli and drugs is of direct clinical and therapeutic relevance to disorders that affect virtually every majo organ system in the body. This proposal is focused primarily on understanding the structure and biophysical properties of the capsaicin- and heat-activated receptor, TRPV1 - perhaps the best-characterized member of the mammalian TRP channel family. Its widely validated role in pain physiology, together with the availability of well characterized pharmacological agents (natural and synthetic), make it a 'poster child' for elucidating basic principles underlying TRP channel pharmacology, structure, and regulation. The studies proposed here are aimed at broadening our understanding of the structural and biophysical principles whereby TRPV1 and related channels are activated or modulated by chemical or physical stimuli. The specific aims are to (i) analyze intrinsic sensitivity of TRPV1 to heat, phospholipids, and other agents in a defined environment consisting of purified channel protein and synthetic lipids; (ii) exploit purified, functional TRPV1 protein for in vitro spectroscopic studies to examine stimulus-evoked conformational movements, and (iii) use voltage-clamp fluorometry to assess the dynamics of stimulus-evoked conformational rearrangements of TRPV1 in cells. Together, these aims will address unresolved issues concerning TRP channel function and structure while laying important groundwork for the long-term goal of obtaining three-dimensional structures of TRPV1 or other TRP channels - which represents a logical and essential next step for the field. Such information is key to the rational development of therapeutic agents that target chronic inflammatory pain syndromes (e.g. arthritis, irritable bowel syndrome, and asthma) and other disorders involving TRP channels.